Moderate exposure to ionizing radiation can be survived with appropriate clinical intervention, which ranges from supportive care to stem cell transplant, depending on dose of exposure. Therefore, rapid triage of exposure victims based on biodosimetry is essential for saving lives and optimizing deployment of resources during a disaster. Current biodosimetry based on parameters such as time to onset of vomiting, lymphocyte depletion kinetics, and the appearance of dicentric chromosomes are less than ideal because they require at least 2-3 days for analysis and they are not easily automated. The goal of this project is to develop a sensitive and automated dosimetric assay. This proposal is significant because development of rapid and automated assays for quantifying exposure would greatly improve triage and thereby improve survival of victims. Our approach is to develop protein-based diagnostics that can be deployed "in the field" without sophisticated technology to assess exposure immediately. Human cells exhibit a robust, radiation-induced proteomic response detectable within minutes following exposure, and many of these proteomic changes show highly reproducible dose- and time-dependence and hence hold great promise for biodosimetry. The objective of this proposal is to test the hypothesis that changes in the human proteome (plasma, urine, or peripheral blood mononuclear cells) in response to radiation can serve as clinical biomarkers to determine radiation exposure levels. This approach is innovative, since there are currently no protein diagnostics for radiation exposure. As an end product to this study, we envision development of blood- or urine-based protein diagnostics, similar to the widely available, over-the-counter pregnancy test. However, rather than monitoring pHCG, our tests would monitor proteins responding to radiation. Specific aim 1: Develop ELISA assays and reagents for quantifying radiation-induced proteomic changes. Specific aim 2: Determine the biodosimetry of radiation-induced protein biomarkers ex vivo (PBMCs) and in vivo in a dog model. Specific aim 3: Determine the biodosimetry of radiation-induced protein biomarkers ex vivo (PBMCs) and in vivo in humans.